大鼠骨髓间充质干细胞分离和培养方法的研究

Study on isolation and culture of rat bone marrow mesenchymal stem cells

目的研究在体外培养和纯化SD大鼠骨髓间充质干细胞(BMSCs)的方法。方法采用改良全骨髓贴壁方法,分离和纯化3～4周的大鼠BMSCs,镜下连续观察细胞的形态变化。流式细胞仪鉴定其表面抗原CD45和CD29的表达情况。结果原代培养的细胞呈圆形和梭形等,24 h后大部分细胞均贴壁。8～10 d可达80%～90%融合,纯化后传代周期为6～8 d。流式细胞术鉴定表明CD45阴性,CD29阳性。结论改良全骨髓贴壁法可有效分离培养大鼠的骨髓间充质干细胞,是一种比较理想的分离培养方法。

Objective To explore a puerification and culture method of bone marrow mesenchymal stem cells（BMSCs） in rat in vitro.Methods BMSCs from bone marrow of 3~4 week old rats were separated and purified by a improved method of whole bone marrow adherent.The morphological changes of BMSCs were observed continuously under the microscope.The expression of surface antigens CD45 and CD29 were analyzed by flow cytometry.Results Cultured primary BMSCs were circular or spindle-shaped and the most BMSCs were adherent within 24 hours,and 80%~90% BMSCs were in fusion within 8~10 days.The Transfer of generation cycle was 6~8 days after purification.The Flow cytometry identification showed that CD29 was positive and CD45 negative.Conclusion The improved method of whole bone marrow adherent.can successfully isolate and culture BMSCs in rats,which is proved to be an optimal methods for cell preparation and culture.